Dynamics of Glucose-induced Membrane Recruitment of Protein Kinase C βII in Living Pancreatic Islet β-Cells

The mechanisms by which glucose may affect protein kinase C (PKC) activity in the pancreatic islet beta-cell are presently unclear. By developing adenovirally expressed chimeras encoding fusion proteins between green fluorescent protein and conventional (betaII), novel (delta), or atypical (zeta) PKCs, we show that glucose selectively alters the subcellular localization of these enzymes dynamically in primary islet and MIN6 beta-cells. Examined by laser scanning confocal or total internal reflection fluorescence microscopy, elevated glucose concentrations induced oscillatory translocations of PKCbetaII to spatially confined regions of the plasma membrane. Suggesting that increases in free cytosolic Ca(2+) concentrations ([Ca(2+)](c)) were primarily responsible, prevention of [Ca(2+)](c) increases with EGTA or diazoxide completely eliminated membrane recruitment, whereas elevation of cytosolic [Ca(2+)](c) with KCl or tolbutamide was highly effective in redistributing PKCbetaII both to the plasma membrane and to the surface of dense core secretory vesicles. By contrast, the distribution of PKCdelta.EGFP, which binds diacylglycerol but not Ca(2+), was unaffected by glucose. Measurement of [Ca(2+)](c) immediately beneath the plasma membrane with a ratiometric "pericam," fused to synaptic vesicle-associated protein-25, revealed that depolarization induced significantly larger increases in [Ca(2+)](c) in this domain. These data demonstrate that nutrient stimulation of beta-cells causes spatially and temporally complex changes in the subcellular localization of PKCbetaII, possibly resulting from the generation of Ca(2+) microdomains. Localized changes in PKCbetaII activity may thus have a role in the spatial control of insulin exocytosis

Exploring gender impact on collaborative care planning: insights from a community mental health service study in Italy

INTRODUCTION: Personal recovery is associated with socio-demographic and clinical factors, and gender seems to influence the recovery process. This study aimed to investigate: i) differences in the recovery goals of men and women users of a community mental health service in Italy; ii) any differences by gender in recovery over six months using the Mental Health Recovery Star (MHRS). METHODS: Service users and staff completed the MHRS together at recruitment and six months later to agree the recovery goals they wished to focus on. Socio-demographic and clinical characteristics and ratings of symptoms (BPRS), needs (CAN), functioning (FPS), and functional autonomy (MPR) were collected at recruitment and six months follow-up. Comparisons between men and women were made using t-tests. RESULTS: Ten women and 15 men completed the MHRS with 19 mental health professionals. Other than gender, men and women had similar socio-demographic, and clinical characteristics at recruitment. Women tended to choose recovery goals that focused on relationships whereas men tended to focus on work related goals. At follow-up, both men and women showed improvement in their recovery (MHRS) and women were less likely to focus on relationship related goals, perhaps because some had found romantic partners. There were also gains for both men and women in engagement with work related activities. Ratings of functional autonomy (MPR) improved for both men and women, and men also showed improvement in symptoms (BPRS) and functioning (FPS). CONCLUSIONS: Our findings suggest that collaborative care planning tools such as the MHRS can assist in identifying individualized recovery goals for men and women with severe mental health problems as part of their rehabilitation

Dense core secretory vesicles revealed as a dynamic Ca2+ store in neuroendocrine cells with a vesicle-associated membrane protein aequorin chimaera

The role of dense core secretory vesicles in the control of cytosolic-free Ca2+ concentrations ([Ca2+]c) in neuronal and neuroendocrine cells is enigmatic. By constructing a vesicle-associated membrane protein 2–synaptobrevin.aequorin chimera, we show that in clonal pancreatic islet β-cells: (a) increases in [Ca2+]c cause a prompt increase in intravesicular-free Ca2+ concentration ([Ca2+]SV), which is mediated by a P-type Ca2+-ATPase distinct from the sarco(endo) plasmic reticulum Ca2+-ATPase, but which may be related to the PMR1/ATP2C1 family of Ca2+ pumps; (b) steady state Ca2+ concentrations are 3–5-fold lower in secretory vesicles than in the endoplasmic reticulum (ER) or Golgi apparatus, suggesting the existence of tightly bound and more rapidly exchanging pools of Ca2+; (c) inositol (1,4,5) trisphosphate has no impact on [Ca2+]SV in intact or permeabilized cells; and (d) ryanodine receptor (RyR) activation with caffeine or 4-chloro-3-ethylphenol in intact cells, or cyclic ADPribose in permeabilized cells, causes a dramatic fall in [Ca2+]SV. Thus, secretory vesicles represent a dynamic Ca2+ store in neuroendocrine cells, whose characteristics are in part distinct from the ER/Golgi apparatus. The presence of RyRs on secretory vesicles suggests that local Ca2+-induced Ca2+ release from vesicles docked at the plasma membrane could participate in triggering exocytosis

Compartmentalized Phosphodiesterase-2 Activity Blunts β-Adrenergic Cardiac Inotropy via an NO/cGMP-Dependent Pathway

β-Adrenergic signaling via cAMP generation and PKA activation mediates the positive inotropic effect of catecholamines on heart cells. Given the large diversity of protein kinase A targets within cardiac cells, a precisely regulated and confined activity of such signaling pathway is essential for specificity of response. Phosphodiesterases (PDEs) are the only route for degrading cAMP and are thus poised to regulate intracellular cAMP gradients. Their spatial confinement to discrete compartments and functional coupling to individual receptors provides an efficient way to control local [cAMP] i in a stimulus-specific manner. By performing real-time imaging of cyclic nucleotides in living ventriculocytes we identify a prominent role of PDE2 in selectively shaping the cAMP response to catecholamines via a pathway involving β 3 -adrenergic receptors, NO generation and cGMP production. In cardiac myocytes, PDE2, being tightly coupled to the pool of adenylyl cyclases activated by β-adrenergic receptor stimulation, coordinates cGMP and cAMP signaling in a novel feedback control loop of the β-adrenergic pathway. In this, activation of β 3 -adrenergic receptors counteracts cAMP generation obtained via stimulation of β 1 /β 2 -adrenoceptors. Our study illustrates the key role of compartmentalized PDE2 in the control of catecholamine-generated cAMP and furthers our understanding of localized cAMP signaling

Years of life that could be saved from prevention of hepatocellular carcinoma

BACKGROUND: Hepatocellular carcinoma (HCC) causes premature death and loss of life expectancy worldwide. Its primary and secondary prevention can result in a significant number of years of life saved. AIM: To assess how many years of life are lost after HCC diagnosis. METHODS: Data from 5346 patients with first HCC diagnosis were used to estimate lifespan and number of years of life lost after tumour onset, using a semi-parametric extrapolation having as reference an age-, sex- and year-of-onset-matched population derived from national life tables. RESULTS: Between 1986 and 2014, HCC lead to an average of 11.5 years-of-life lost for each patient. The youngest age-quartile group (18-61 years) had the highest number of years-of-life lost, representing approximately 41% of the overall benefit obtainable from prevention. Advancements in HCC management have progressively reduced the number of years-of-life lost from 12.6 years in 1986-1999, to 10.7 in 2000-2006 and 7.4 years in 2007-2014. Currently, an HCC diagnosis when a single tumour <2 cm results in 3.7 years-of-life lost while the diagnosis when a single tumour 65 2 cm or 2/3 nodules still within the Milan criteria, results in 5.0 years-of-life lost, representing the loss of only approximately 5.5% and 7.2%, respectively, of the entire lifespan from birth. CONCLUSIONS: Hepatocellular carcinoma occurrence results in the loss of a considerable number of years-of-life, especially for younger patients. In recent years, the increased possibility of effectively treating this tumour has improved life expectancy, thus reducing years-of-life lost